Qualitative and Quantitative Analysis of Proteins in Milk

Cow’s milk is an important source of protein in human nutrition. On average, cow’s milk contains 3.5% protein, which can be classified as either casein or whey. The composition of milk proteins varies considerably due to breed, stage of lactation, health and nutritional status of the animal. Casein proteins are the primary components in cheese, whereas whey proteins are by-products of cheesemaking. Recognition of the nutritional, functional, and bioactive properties of whey proteins and their derivatives has been gradually established, rendering them an increasingly valued ingredient in traditional and new foods, including functional foods. Characterization and quantification of individual whey proteins, like β-lactoglobulin, α-lactalbumin, and lactoferrin, are of crucial importance to industrial dairy processes. Labor intensive wet-chemical methods are still being used for this purpose, but a rapid quantification method for individual whey proteins is highly desired. This thesis details studies to use Fourier transform mid-infrared (MIR) spectroscopy for quantification of the two major whey proteins, β-lactoglobulin and α-lactalbumin, and Fourier transform near-infrared (NIR) spectroscopy for quantification of the minor whey protein, lactoferrin, in aqueous whey solutions. In the first study, MIR spectra of whey samples were used to develop a support vector regression (SVR) prediction model to quantify β-lactoglobulin and α-lactalbumin, achieving R² values of 92.8% for β-lactoglobulin and 92.7% for α-lactalbumin. The second study utilized NIR spectroscopy to quantify the low abundance, high-value protein, lactoferrin. Quantification of lactoferrin was based on high performance liquid chromatography with diode array detection (HPLC-DAD) calibration curves for commercial lactoferrin that were detected at 220 nm. Linearity of the calibration curve with seven different concentrations of lactoferrin between 0.02-2.0 mg mL-1, triplicate analysis had an R2 value of 0.989. The concentration of lactoferrin from the whey portion of a Cornell University obtained reference sample and whey from skim milk were determined to be within the range 0.01-0.10 mg mL-1. These data were combined with NIR spectra of aqueous whey to generate a data library for chemometric model development.